DESCRIPTION: The long-term objective of this project is to provide a thorough understanding of how the low-density lipoprotein receptor (LDLR) folds into its native structure and recognizes its lipoprotein ligands. The LDLR is the primary mechanism for uptake of plasma cholesterol into cells. When the LDLR is unable to clear cholesterol-containing lipoproteins sufficiently from the blood, an elevated plasma cholesterol level results. A high plasma cholesterol level is a major risk for heart disease, the leading cause of death in the United States. Over 150 different mutations of the LDLR give rise to familial hypercholesterolemia (FH), which is characterized clinically by an elevated concentration of plasma LDL and cholesterol. Detailed structural and biochemical studies of LDLR-lipoprotein interactions have been elusive, because the receptor protein is large and membrane bound. However, the ligand-binding domain of the LDLR is composed of a series of autonomously structured, non-identical tandem repeats that can be studied in isolation from the rest of the receptor. In previous work, the PI has shown that a critical repeat (repeat 5) within the ligand-binding domain of the receptor can be folded to its native structure after expression in bacteria, and that calcium is required for proper folding of this domain. During the period of grant support, he plans (1) to determine the principles that govern proper folding of this prototypic repeat of the LDLR ligand-binding domain into its native structure, and (2) to elucidate the detailed molecular basis for ligand-binding by the LDLR, relying on the folding studies to identify potential sites of receptor-ligand interaction. This work will have broad implications for the mechanism of ligand interactions. This work will have broad implications for the mechanism of ligand recognition by the wide variety of proteins that contain structural motifs homologous to those found in the ligand-binding domain of the LDLR, including proteins implicated in G-protein couple signaling, brain development, and the immune response. Understanding the basis for ligand recognition by LDL-A repeats may ultimately allow the alteration of the ligand-binding properties of LDL-A repeats to create novel receptors for arbitrary target ligands. Eventually, small molecules may be identified which suppress the folding defects in some of the FH mutations, and which might serve as therapeutics for patients with FH.